Vascular remodeling, a common process in many vascular diseases, describes morphological changes within the vascular wall and adjacent perivascular structures Inflammation, endothelial dysfunction, and vascular smooth muscle cell migration and proliferation are the main factors triggering this remodeling process, which may lead to neointimal hyperplasia, medial dysplasia, perivascular thickening, or fibrosis and vascular calcification 45. Vascular remodeling also occurs after interventional procedures such as angioplasty and vascular stenting. Veins are frequently used in coronary artery bypass grafting, which may fail in the short term because of acute graft failure due to thrombosis or in the long term due to pathological vascular remodeling, a complex process that is poorly understood despite its clinical relevance for other diseases. A better understanding of the molecular signaling networks and cellular responses holds tremendous potential for patients undergoing these procedures. We have embarked on a series of investigations to determine cellular responses and local signaling pathways in a murine vein graft model. Project 1: AD-HIES clinical features and signaling pathways that we identified as those affected in AD-HIES fibroblasts point to angiogenesis defects as factors that contribute to AD-HIES pathologies. Therefore, we used co-culture in vitro angiogenesis assays to determine the ability of AD-HIES fibroblasts to support angiogenesis. Our studies confirmed that AD-HIES fibroblasts have decreased ability to promote tube formation by HUVEC cells. To investigate the ability of AD-HIES fibroblasts to support angiogenesis in vivo, we are using a murine model of hind-limb ischemia, in which surgical ligation of the femoral artery at specific site leads to arteriogenesis in femoral collaterals and angiogenesis in distal ischemic muscles.42 Preliminary experiments demonstrated that injecting normal fibroblasts into the calf muscle improves restoration of blood flow while injecting AD-HIES fibroblasts does not. These results indicate that AD-HIES fibroblasts are deficient in supporting angiogenesis in vivo. Project 2: Veins grafted into an arterial environment undergo complex vascular remodeling. EndMT is the process by which endothelial cells lose their cell-specific markers and morphology and acquire a mesenchymal cell-like phenotype. However, the role of EndMT and the mechanisms regulating cell phenotype adaptation during human vein graft remodeling are poorly understood. Using two independent endothelial lineage tracing systems, we demonstrated that, at 35 days after vein grafting, half the cells populating the neointima were of endothelial origin. These endothelial-derived cells had lost their endothelial phenotype and acquired smooth muscle cell-like properties, yet they failed to develop a fully mature smooth muscle cell phenotype. We detected early activation of the TGF-b-Smad2/3-Slug signaling pathway and confirmed that TGF-b-Smad2/3-Slug signaling pathway-mediated EndMT plays a pivotal role in regulating vein graft remodeling. We are further investigating BMP9/Smad1-mediated endothelial cell survival and its impact on acute vein graft patency and long-term outcome in vein graft remodeling due to the development of neointimal hyperplasia, but also in thrombosis formation leading to early graft failure. We hypothesize that activating the BMP9/Smad1 pathway positively regulates endothelial function to benefit murine vein grafting.